Atrial (ANP) and B-type natriuretic peptides (BNP) modulate blood pressure and volume through the stimulation of cyclic GMP production by their guanylyl cyclase-A (GC-A) receptor. We identified a novel isoform of GC-A which is the result of differential splicing of exon 4. The deletion of a 51-bp sequence is predicted to delete 17 amino acids (Lys314–Gln330) in the membrane-distal part of the extracellular domain. Reverse transcription-PCR analyses demonstrated low messenger RNA expression levels of spliced GC-A in all tissues. Homology modeling suggested that the alterations in the protein structure could interfere with ANP binding or signaling. Indeed, functional studies in transfected HEK 293 cells demonstrated that binding of ANP and ANP-induced cyclic GMP formation by GC-ADeltaLys314-Gln330 were totally abolished. Furthermore, cotransfection studies showed that this GC-A variant forms heterodimers with the wild type receptor and inhibits ligand-inducible cGMP generation. Lastly we examined whether enhanced alternative splicing could contribute to the known heterologous and/or homologus desensitization of GC-A. Treatment of mice with angiotensin II (300 ng/kg/min during 7 days) resulted in enhanced pulmonary mRNA expression of spliced GC-A, which was concomitant to diminished GC-A/cGMP responses to ANP. Also treatment of cultured murine lung endothelial cells with ANP (100 nM during 7h) resulted in enhanced mRNA expression of the GC-A isoform. We conclude that alternative splicing can regulate endogenous ANP/GC-A signaling. Our observations in vivo/in vitro indicate that Angiotensin II- and ANP-induced alternative splicing of GC-A may represent a novel mechanism for reducing the sensitivity of the receptor to ANP.

Supported bei Deutsche Forschungsgemeinschaft SFB 487